Field of the Invention
The present invention relates to methods for producing a morpholino oligonucleotide. The present invention also relates to morpholino nucleotides used as a starting material in such a production method.
Discussion of the Background
Morpholino oligonucleotides are compounds attracting attention for their use as antisense oligonucleotides, since they show high affinity for DNA and RNA, resistance to various nucleases, stability in vivo, and low toxicity (see Summerton, J. et al., Antisense and Nucleic Acid Drug Development, 1997, Vol. 7, p. 187, which is incorporated herein by reference in its entirety).
As a production method of morpholino oligonucleotides, a solid-phase synthesis and a liquid-phase synthesis have been reported (see Harakawa et al., Bioorganic & Medicinal Chemistry Letters, 2012, Vol. 22, p. 1445-1447; WO 91/09033; WO 2008/008113; US 2009/0131632 A1; WO 2009/064471; and WO 2012/043730, all of which are incorporated herein by reference in their entireties).
Solid-phase synthesis is advantageous from the aspect of speed since it enables automatic synthesis. On the other hand, it is not suitable for industrial large scale synthesis since scaling-up is limited due to facility restriction, and low reactivity requires use of an excess monomer to be the reagent in a nucleotide elongation reaction. Also, it is associated with defects in that confirmation of the progress status of the reaction in an intermediate stage, analysis of intermediate structure and the like are difficult.
On the other hand, liquid-phase synthesis has problems of solubility, complexity of work-up and the like, and a large-scale and rapid synthesis of a morpholino oligonucleotide having a chain length utilizable as an antisense pharmaceutical product has been difficult.
Thus, there remains a need for improved methods for producing a morpholino oligonucleotide.